Acupuncture is an ancient therapeutic modality whose mechanisms of action only recently are being defined. While clinical trials are defining efficacy in areas such as pain, infections and cardiovascular disease, including hypertension, myocardial ischemia and arrhythmias, it is clear that we know little about its mechanisms of action. Over the last three year, this grant has explored the function of a long-loop pathway, including the arcuate nucleus (ARC) in the ventral hypothalamus, the ventrolateral periaqueductal gray (vIPAG) in the midbrain and the rostral ventrolateral medulla (rVLM) in regulation of visceral sympathoexcitatory reflexes. We have demonstrated excitatory input from acupuncture points located along the distal median and deep peroneal somatic nerves activated during electroacupuncture (EA) to the ARC, vIPAG and rVLM. The ARC provides an excitatory projection to the vIPAG, which in turn, provides an inhibitory projection to the rVLM that ultimately regulates sympathetic premotor neurons that innervate the spinal cord sympathetic outflow to the cardiovascular system. The present grant refines and extends our previous observations with three specific aims. The first aim evaluates the significance of reciprocal projections between the ARC and the vIPAG, which we hypothesize, reinforces input in the ARC from somatic afferent stimulation during EA. The second aim identifies the role of specific neurotransmitter systems, including glutamate and its ionotropic receptors in ARC and vIPAG, acetylcholine in ARC and the endocannabinoid system in vIPAG, which we hypothesize, inhibits the release of gamma-aminobutyric acid to disinhibit vIPAG neuronal activity and ultimately potentiate vIPAG inhibition of the rVLM. The third aim examines the role of midline medullary nuclei, especially the nuclei raphe obscurus, magnus and pallidus, which we believe receive excitatory input from the vIPAG and, in turn, inhibit rVLM activity. Studies will use a combination of anatomical, cellular electrophysiological, pharmacological, microcollection-microassay and whole animal reflex approaches to test these hypotheses. This added knowledge will provide physicians and scientists with a greater mechanistic understanding of EA's cardiovascular influence, which will increase its clinical acceptance, e.g., in patients with cardiovascular disease. Furthermore, knowledge of the mechanistic basis of acupuncture may allow practitioners to optimize the clinical cardiovascular responses to treatment.